Pore formation by the CDTb component of the Clostridioides difficile binary toxin is Ca2+-dependent
Abstract Clostridioides difficile infection (CDI) is one of the five most urgent bacterial threats in the United States. Furthermore, hypervirulent CDI strains express a third toxin termed the C. difficile binary toxin (CDT), and its molecular mechanism for entering host cells is not fully elucidate...
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Nature Portfolio
2025-06-01
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| Series: | Communications Biology |
| Online Access: | https://doi.org/10.1038/s42003-025-08343-x |
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| author | Dinendra L. Abeyawardhane Spiridon E. Sevdalis Kaylin A. Adipietro Raquel Godoy-Ruiz Kristen M. Varney Izza F. Nawaz Alejandro X. Spittel Daniel Hunter Richard R. Rustandi Vitalii I. Silin Amedee des Georges Mary E. Cook Edwin Pozharski David J. Weber |
| author_facet | Dinendra L. Abeyawardhane Spiridon E. Sevdalis Kaylin A. Adipietro Raquel Godoy-Ruiz Kristen M. Varney Izza F. Nawaz Alejandro X. Spittel Daniel Hunter Richard R. Rustandi Vitalii I. Silin Amedee des Georges Mary E. Cook Edwin Pozharski David J. Weber |
| author_sort | Dinendra L. Abeyawardhane |
| collection | DOAJ |
| description | Abstract Clostridioides difficile infection (CDI) is one of the five most urgent bacterial threats in the United States. Furthermore, hypervirulent CDI strains express a third toxin termed the C. difficile binary toxin (CDT), and its molecular mechanism for entering host cells is not fully elucidated. Like other AB-type binary toxins, CDT enters host cells via endosomes. Here we show via surface plasmon resonance and electrochemical impedance spectroscopy that the cell-binding component of CDT, termed CDTb, binds and form pores in lipid bilayers in the absence of its enzymatic component, CDTa. This occurs upon lowering free Ca2+ ion concentration, and not by decreasing pH, as found for other binary toxins (i.e., anthrax). Cryogenic electron microscopy (CryoEM), X-ray crystallography, and nuclear magnetic resonance (NMR) studies show that dissociation of Ca2+ from a single site in receptor binding domain 1 (RBD1) of CDTb triggers conformational exchange in CDTb. These and structure/function studies of a Ca2+-binding double mutant targeting RBD1 (i.e., D623A/D734A) support a model in which dissociation of Ca2+ from RBD1 induces dynamic properties in CDTb that enable it to bind and form pores in lipid bilayers. |
| format | Article |
| id | doaj-art-d8d61c8064ea4fc6be04570da3fa8b2b |
| institution | OA Journals |
| issn | 2399-3642 |
| language | English |
| publishDate | 2025-06-01 |
| publisher | Nature Portfolio |
| record_format | Article |
| series | Communications Biology |
| spelling | doaj-art-d8d61c8064ea4fc6be04570da3fa8b2b2025-08-20T02:07:45ZengNature PortfolioCommunications Biology2399-36422025-06-018111410.1038/s42003-025-08343-xPore formation by the CDTb component of the Clostridioides difficile binary toxin is Ca2+-dependentDinendra L. Abeyawardhane0Spiridon E. Sevdalis1Kaylin A. Adipietro2Raquel Godoy-Ruiz3Kristen M. Varney4Izza F. Nawaz5Alejandro X. Spittel6Daniel Hunter7Richard R. Rustandi8Vitalii I. Silin9Amedee des Georges10Mary E. Cook11Edwin Pozharski12David J. Weber13Department of Biochemistry and Molecular Biology, University of Maryland School of MedicineDepartment of Biochemistry and Molecular Biology, University of Maryland School of MedicineDepartment of Biochemistry and Molecular Biology, University of Maryland School of MedicineDepartment of Biochemistry and Molecular Biology, University of Maryland School of MedicineDepartment of Biochemistry and Molecular Biology, University of Maryland School of MedicineDepartment of Biochemistry and Molecular Biology, University of Maryland School of MedicineDepartment of Biochemistry and Molecular Biology, University of Maryland School of MedicineDepartment of Biochemistry and Molecular Biology, University of Maryland School of MedicineMerck & Co. Inc.Institute for Bioscience and Biotechnology Research (IBBR), University of MarylandStructural Biology Initiative, CUNY Advanced Science Research CenterDepartment of Biochemistry and Molecular Biology, University of Maryland School of MedicineDepartment of Biochemistry and Molecular Biology, University of Maryland School of MedicineDepartment of Biochemistry and Molecular Biology, University of Maryland School of MedicineAbstract Clostridioides difficile infection (CDI) is one of the five most urgent bacterial threats in the United States. Furthermore, hypervirulent CDI strains express a third toxin termed the C. difficile binary toxin (CDT), and its molecular mechanism for entering host cells is not fully elucidated. Like other AB-type binary toxins, CDT enters host cells via endosomes. Here we show via surface plasmon resonance and electrochemical impedance spectroscopy that the cell-binding component of CDT, termed CDTb, binds and form pores in lipid bilayers in the absence of its enzymatic component, CDTa. This occurs upon lowering free Ca2+ ion concentration, and not by decreasing pH, as found for other binary toxins (i.e., anthrax). Cryogenic electron microscopy (CryoEM), X-ray crystallography, and nuclear magnetic resonance (NMR) studies show that dissociation of Ca2+ from a single site in receptor binding domain 1 (RBD1) of CDTb triggers conformational exchange in CDTb. These and structure/function studies of a Ca2+-binding double mutant targeting RBD1 (i.e., D623A/D734A) support a model in which dissociation of Ca2+ from RBD1 induces dynamic properties in CDTb that enable it to bind and form pores in lipid bilayers.https://doi.org/10.1038/s42003-025-08343-x |
| spellingShingle | Dinendra L. Abeyawardhane Spiridon E. Sevdalis Kaylin A. Adipietro Raquel Godoy-Ruiz Kristen M. Varney Izza F. Nawaz Alejandro X. Spittel Daniel Hunter Richard R. Rustandi Vitalii I. Silin Amedee des Georges Mary E. Cook Edwin Pozharski David J. Weber Pore formation by the CDTb component of the Clostridioides difficile binary toxin is Ca2+-dependent Communications Biology |
| title | Pore formation by the CDTb component of the Clostridioides difficile binary toxin is Ca2+-dependent |
| title_full | Pore formation by the CDTb component of the Clostridioides difficile binary toxin is Ca2+-dependent |
| title_fullStr | Pore formation by the CDTb component of the Clostridioides difficile binary toxin is Ca2+-dependent |
| title_full_unstemmed | Pore formation by the CDTb component of the Clostridioides difficile binary toxin is Ca2+-dependent |
| title_short | Pore formation by the CDTb component of the Clostridioides difficile binary toxin is Ca2+-dependent |
| title_sort | pore formation by the cdtb component of the clostridioides difficile binary toxin is ca2 dependent |
| url | https://doi.org/10.1038/s42003-025-08343-x |
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